Showing papers by "Baylor College of Medicine published in 1992"

Mice deficient for p53 are developmentally normal but susceptible to spontaneous tumours

Abstract: Mutations in the p53 tumour-suppressor gene are the most frequently observed genetic lesions in human cancers. To investigate the role of the p53 gene in mammalian development and tumorigenesis, a null mutation was introduced into the gene by homologous recombination in murine embryonic stem cells. Mice homozygous for the null allele appear normal but are prone to the spontaneous development of a variety of neoplasms by 6 months of age. These observations indicate that a normal p53 gene is dispensable for embryonic development, that its absence predisposes the animal to neoplastic disease, and that an oncogenic mutant form of p53 is not obligatory for the genesis of many types of tumours.

Methylation of HpaII and HhaI sites near the polymorphic CAG repeat in the human androgen-receptor gene correlates with X chromosome inactivation.

Abstract: The human androgen-receptor gene (HUMARA; GenBank) contains a highly polymorphic trinucleotide repeat in the first exon. We have found that the methylation of HpaII and HhaI sites less than 100 bp away from this polymorphic short tandem repeat (STR) correlates with X inactivation. The close proximity of the restriction-enzyme sites to the STR allows the development of a PCR assay that distinguishes between the maternal and paternal alleles and identifies their methylation status. The accuracy of this assay was tested on (a) DNA from hamster/human hybrid cell lines containing either an active or inactive human X chromosome; (b) DNA from normal males and females; and (c) DNA from females showing nonrandom patterns of X inactivation. Data obtained using this assay correlated substantially with those obtained using the PGK, HPRT, and M27 beta probes, which detect X inactivation patterns by Southern blot analysis. In order to demonstrate one application of this assay, we examined X inactivation patterns in the B lymphocytes of potential and obligate carriers of X-linked agammaglobulinemia.

An unstable triplet repeat in a gene related to myotonic muscular dystrophy.

Abstract: Synthetic oligonucleotides containing GC-rich triplet sequences were used in a scanning strategy to identify unstable genetic sequences at the myotonic dystrophy (DM) locus. A highly polymorphic GCT repeat was identified and found to be unstable, with an increased number of repeats occurring in DM patients. In the case of severe congenital DM, the paternal triplet allele was inherited unaltered while the maternal, DM-associated allele was unstable. These studies suggest that the mutational mechanism leading to DM is triplet amplification, similar to that occurring in the fragile X syndrome. The triplet repeat sequence is within a gene (to be referred to as myotonin-protein kinase), which has a sequence similar to protein kinases.

Mice deficient for Rb are nonviable and show defects in neurogenesis and haematopoiesis.

Abstract: The retinoblastoma gene, a prototypic tumour-suppressor gene, encodes a nuclear phosphoprotein (Rb). To understand better the role of Rb in development and in tumorigenesis, mice with an insertional mutation in exon 20 of the Rb-1 locus were generated. Homozygous mutants die before the 16th embryonic day with multiple defects. The haematopoietic system is abnormal; there is a significant increase in the number of immature nucleated erythrocytes. In the nervous system, ectopic mitoses and massive cell death are found, particularly in the hindbrain. All spinal ganglion cells die, but the neural retina is unaffected. Transfer of the human retinoblastoma (RB) mini-transgene into the mutant mice corrects the developmental defects. Thus, Rb is essential for normal mouse development.

Early enteral feeding, compared with parenteral, reduces postoperative septic complications. The results of a meta-analysis.

Abstract: This two-part meta-analysis combined data from eight prospective randomized trials designed to compare the nutritional efficacy of early enteral (TEN) and parenteral (TPN) nutrition in high-risk surgical patients. The combined data gave sufficient patient numbers (TEN, n = 118; TPN, n = 112) to adequately address whether route of substrate delivery affected septic complication incidence. Phase I (dropouts excluded) meta-analysis confirmed data homogeneity across study sites, that TEN and TPN groups were comparable, and that significantly fewer TEN patients experienced septic complications (TEN, 18%; TPN, 35%; p = 0.01). Phase II meta-analysis, an intent-to-treat analysis (dropouts included), confirmed that fewer TEN patients developed septic complications. Further breakdown by patient type showed that all trauma and blunt trauma subgroups had the most significant reduction in septic complications when fed enterally. In conclusion, this meta-analysis attests to the feasibility of early postoperative TEN in high-risk surgical patients and that these patients have reduced septic morbidity rates compared with those administered TPN.

Impaired Long-Term Potentiation, Spatial Learning, and Hippocampal Development in fyn Mutant Mice

Abstract: Mice with mutations in four nonreceptor tyrosine kinase genes, fyn, src, yes, and abl, were used to study the role of these kinases in long-term potentiation (LTP) and in the relation of LTP to spatial learning and memory. All four kinases were expressed in the hippocampus. Mutations in src, yes, and abl did not interfere with either the induction or the maintenance of LTP. However, in fyn mutants, LTP was blunted even though synaptic transmission and two short-term forms of synaptic plasticity, paired-pulse facilitation and post-tetanic potentiation, were normal. In parallel with the blunting of LTP, fyn mutants showed impaired spatial learning, consistent with a functional link between LTP and learning. Although fyn is expressed at mature synapses, its lack of expression during development resulted in an increased number of granule cells in the dentate gyrus and of pyramidal cells in the CA3 region. Thus, a common tyrosine kinase pathway may regulate the growth of neurons in the developing hippocampus and the strength of synaptic plasticity in the mature hippocampus.

Target enzyme recognition by calmodulin: 2.4 A structure of a calmodulin-peptide complex

Abstract: The crystal structure of calcium-bound calmodulin (Ca(2+)-CaM) bound to a peptide analog of the CaM-binding region of chicken smooth muscle myosin light chain kinase has been determined and refined to a resolution of 2.4 angstroms (A). The structure is compact and has the shape of an ellipsoid (axial ratio approximately 2:1). The bound CaM forms a tunnel diagonal to its long axis that engulfs the helical peptide, with the hydrophobic regions of CaM melded into a single area that closely covers the hydrophobic side of the peptide. There is a remarkably high pseudo-twofold symmetry between the closely associated domains. The central helix of the native CaM is unwound and expanded into a bend between residues 73 and 77. About 185 contacts (less than 4 A) are formed between CaM and the peptide, with van der Waals contacts comprising approximately 80% of this total.

Alpha-inhibin is a tumour-suppressor gene with gonadal specificity in mice.

Abstract: The inhibins are α:β heterodimeric growth factors that are members of the transforming growth factor-β family To understand the physiological roles of the inhibins in mammalian development and reproduction, a targeted deletion of the α-inhibin gene was generated by homologous recombination in mouse embryonic stem cells Mice homozygous for the null allele (inhibin-deficient) initially develop normally but every mouse ultimately develops mixed or incompletely differentiated gonadal stromal tumours either unilaterally or bilaterally Inhibin is thus a critical negative regulator of gonadal stromal cell proliferation and the first secreted protein identified to have tumour-suppressor activity

Effect of treatment of helicobacter pylori infection on the long-term recurrence of gastric or duodenal ulcer: a randomized, controlled study

Abstract: OBJECTIVE To determine the effect of treating Helicobacter pylori infection on the recurrence of gastric and duodenal ulcer disease. DESIGN Follow-up of up to 2 years in patients with healed ulcers who had participated in randomized, controlled trials. SETTING A Veterans Affairs hospital. PARTICIPANTS A total of 109 patients infected with H. pylori who had a recently healed duodenal (83 patients) or gastric ulcer (26 patients) as confirmed by endoscopy. INTERVENTION Patients received ranitidine, 300 mg, or ranitidine plus triple therapy. Triple therapy consisted of tetracycline, 2 g; metronidazole, 750 mg; and bismuth subsalicylate, 5 or 8 tablets (151 mg bismuth per tablet) and was administered for the first 2 weeks of treatment; ranitidine therapy was continued until the ulcer had healed or 16 weeks had elapsed. After ulcer healing, no maintenance antiulcer therapy was given. MEASUREMENTS Endoscopy to assess ulcer recurrence was done at 3-month intervals or when a patient developed symptoms, for a maximum of 2 years. RESULTS The probability of recurrence for patients who received triple therapy plus ranitidine was significantly lower than that for patients who received ranitidine alone: for patients with duodenal ulcer, 12% (95% CI, 1% to 24%) compared with 95% (CI, 84% to 100%); for patients with gastric ulcer, 13% (CI, 4% to 31%) compared with 74% (44% to 100%). Fifty percent of patients who received ranitidine alone for healing of duodenal or gastric ulcer had a relapse within 12 weeks of healing. Ulcer recurrence in the triple therapy group was related to the failure to eradicate H. pylori and to the use of nonsteroidal anti-inflammatory drugs. CONCLUSIONS Eradication of H. pylori infection markedly changes the natural history of peptic ulcer in patients with duodenal or gastric ulcer. Most peptic ulcers associated with H. pylori infection are curable.

Expression, self-assembly, and antigenicity of the Norwalk virus capsid protein.

Abstract: Norwalk virus capsid protein was produced by expression of the second and third open reading frames of the Norwalk virus genome, using a cell-free translation system and baculovirus recombinants. Analysis of the expressed products showed that the second open reading frame encodes a protein with an apparent molecular weight of 58,000 (58K protein) and that this protein self-assembles to form empty viruslike particles similar to native capsids in size and appearance. The antigenicity of these particles was demonstrated by immunoprecipitation and enzyme-linked immunosorbent assays of paired serum samples from volunteers who developed illness following Norwalk virus challenge. These particles also induced high levels of Norwalk virus-specific serum antibody in laboratory animals following parenteral inoculation. A minor 34K protein was also found in infected insect cells. Amino acid sequence analysis of the N terminus of the 34K protein indicated that the 34K protein was a cleavage product of the 58K protein. The availability of large amounts of recombinant Norwalk virus particles will allow the development of rapid, sensitive, and reliable tests for the diagnosis of Norwalk virus infection as well as the implementation of structural studies.

The Declining Risk of Post-Transfusion Hepatitis C Virus Infection

Abstract: Background. The most common serious complication of blood transfusion is post-transfusion hepatitis from the hepatitis C virus (HCV). Blood banks now screen blood donors for surrogate markers of non-A, non-B hepatitis and antibodies to HCV, but the current risk of post-transfusion hepatitis C is unknown. Methods. From 1985 through 1991, blood samples and medical information were obtained prospectively from patients before and at least six months after cardiac surgery. The stored serum samples were tested for antibodies to HCV by enzyme immunoassay, and by recombinant immunoblotting if positive. Results. Of the 912 patients who received transfusions before donors were screened for surrogate markers, 35 seroconverted to HCV, for a risk of 3.84 percent per patient (0.45 percent per unit transfused). For the 976 patients who received transfusions after October 1986 with blood screened for surrogate markers, the risk of seroconversion was 1.54 percent per patient (0.19 percent per unit). For the 522 p...

Aging of the human retina. Differential loss of neurons and retinal pigment epithelial cells.

Abstract: The impact of aging on cell loss in the human retina was examined in foveal and temporal equatorial regions in eyes from 35 donors with ages spanning a 78-yr period from the second to the ninth decade of life. Equatorial cones and retinal pigment epithelial cells (RPE) decreased at uniform rates from the second to the ninth decade, 16 and 14 cells/mm2/yr, respectively. Equatorial rods and cells in the ganglion cell layer (GCL) showed nonuniform rate decreases with age. The rates of rod and GCL cell loss were faster between the second and fourth decades (970 and 9 cells/mm2/yr, respectively) than between the fourth and ninth decades (570-330 and 6-3 cells/mm2/yr). The rod and GCL cell densities at the temporal equator maintained a constant ratio (rods-GCL cell ratio = 103 +/- 0.4, mean +/- standard deviation) and the same reduction slope ratio at different times during aging. Thus, the equatorial rod and GCL cell losses were correlated statistically. The ratio of equatorial photoreceptors to RPE cells showed no significant change with age, suggesting parallel loss of these closely apposed cells. At the foveal center, the variability of cone density between individuals in each decade grouping was large (1.7- to threefold). No significant differences were found in cone or RPE cell densities at the foveal center from the second to ninth decade, suggesting that the densities of foveal cones and RPE cells were stable throughout this period. Foveal RPE density was significantly higher than equatorial RPE density in each age group. No significant difference was found between the equatorial photoreceptor-RPE ratio and foveal cone-RPE ratio in any age group. Cells in the GCL in the fovea decreased by approximately 16% from the second to the sixth decade. These results indicated that (1) rod photoreceptors and cells in the GCL were more vulnerable to loss during aging than cones; (2) photoreceptors and RPE cells showed parallel changes during aging; and (3) the photoreceptor loss accompanying aging was less pronounced in the fovea than in the peripheral retina.

Visual response latencies in striate cortex of the macaque monkey

Abstract: 1 Many lines of evidence suggest that signals relayed by the magnocellular and parvocellular subdivisions of the primate lateral geniculate nucleus (LGN) maintain their segregation in cortical processing We have examined two response properties of units in the striate cortex of macaque monkeys, latency and transience, with the goal of assessing whether they might be used to infer specific geniculate contributions Recordings were made from 298 isolated units and 1,129 multiunit sites in the striate cortex in four monkeys Excitotoxin lesions that selectively affected one or the other LGN subdivision were made in three animals to demonstrate directly the magnocellular and parvocellular contributions An additional 435 single units and 551 multiunit sites were recorded after the ablations 2 Most units in striate cortex had visual response latencies in the range of 30-50 ms under the stimulus conditions used The earliest neuronal responses in striate cortex differed appreciably between individuals The shortest latency recorded in the four animals ranged from 20 to 31 ms Comparable values were obtained from both single unit and multiunit sites After lesions were made in the magnocellular subdivision of the LGN in two animals, the shortest response latencies were 7 and 10 ms later than before the ablations A larger lesion in the parvocellular subdivision of another animal produced no such shift Thus it appears that the first 7-10 ms of cortical activation can be attributed to activation relayed by the magnocellular layers of the LGN 3 The units with the shortest latencies were all found in layers 4C or 6 and their responses were among the most transient in striate cortex Furthermore, their responses all showed a pronounced periodicity at a frequency of 50-100 Hz This periodicity was stimulus locked, and the responses of all short-latency units oscillated in phase 4 An index of response transience was computed for the units recorded in striate cortex The distribution of this index was unimodal and gave no suggestion of distinct contributions from the geniculate subdivisions Magnocellular and the parvocellular lesions affected the overall transience of responses in striate cortex The changes, however, were very small; extremely transient responses and extremely sustained responses survived both types of lesions 5 A characteristic profile was observed in the response latencies in superficial layers Latencies appeared to increase monotonically from layer 4 toward the surface of cortex, with the most superficial neurons not becoming active until 15 ms after responses were observed in layer 4C(ABSTRACT TRUNCATED AT 400 WORDS)

EMG analysis of the scapular muscles during a shoulder rehabilitation program

Abstract: The purpose of this study was to determine which exercises most effectively use the scapular muscles. Eight muscles in 9 healthy subjects were studied with indwelling electromyographic electrodes and cinema tography while performing 16 exercises. The 8 muscles studied were the upper, middle, and lower trapezius; levator scapula; rhomboids; pectoralis minor; and the middle and lower serratus anterior. Each exercise was divided into arcs of motion and the electromyographic activity was quantified as a percentage of the maximal manual muscle test. The optimal exercises for each muscle were identified based on intensity (greater than 50% maximal manual muscle test) and duration (over at least 3 consecutive arcs of motion) of the muscle activity. Twelve of the exercises qualified as top exer cises for all of the muscles. On further analysis, a group of 4 exercises was shown to make up the core of a scapular muscle strengthening program. Those 4 ex ercises include scaption (scapular plane elevation), row ing, p...

Abbreviated laparotomy and planned reoperation for critically injured patients.

Abstract: The triad of hypothermia, acidosis, and coagulopathy in critically injured patients is a vicious cycle that, if uninterrupted, is rapidly fatal. During the past 7.5 years, 200 patients were treated with unorthodox techniques to abruptly terminate the laparotomy and break the cycle. One hundred seventy patients (85%) suffered penetrating injuries and 30 (15%) were victims of blunt trauma. The mean Revised Trauma Score, Injury Severity Score, and Trauma Index Severity Score age combination index predicted survival were 5.06%, 33.2%, and 57%, respectively. Resuscitative thoracotomies were performed in 60 (30%) patients. After major sources of hemorrhage were controlled, the following clinical and laboratory mean values were observed: red cell transfusions--22 units, core temperature--32.1 C, and pH--7.09. Techniques to abbreviate the operation included the ligation of enteric injuries in 34 patients, retained vascular clamps in 13, temporary intravascular shunts in four, packing of diffusely bleeding surfaces in 171, and the use of multiple towel clips to close only the skin of the abdominal wall in 178. Patients then were transported to the surgical intensive care unit for vigorous correction of metabolic derangements and coagulopathies. Ninety-eight patients (49%) survived to undergo planned reoperation (mean delay 48.1 hours), and 66 of 98 (67%) survived to leave the hospital. With the exception of intravascular shunts, there were survivors who were treated by each of the unorthodox techniques. Of 102 patients who died before reoperation 68 (67%) did so within 2 hours of the initial procedure. Logistic regression showed that red cell transfusion rate and pH may be helpful in determining when to consider abbreviated laparotomy. The authors conclude that patients with hypothermia, acidosis, and coagulopathy are at high risk for imminent death, and that prompt termination of laparotomy with the use of the above techniques is a rational approach to an apparently hopeless situation.

Molecular cloning of the receptor for human antidiuretic hormone.

Abstract: ANTIDIURESIS, the recovery of water from the lumen of the renal collecting tubule, is regulated by the hypothalamic release of antidiuretic hormone (ADH), which binds to specific receptors on renal collecting tubule cells, stimulates adenylyl cyclase and promotes the cyclic AMP-mediated incorporation of water pores into the luminal surface of these cells'1–3. We report here the isolation of the human ADH receptor gene using a genomic expression cloning approach4. The gene was used to clone the complementary DNA from a human renal library. The deduced amino-acid sequence of the receptor yields a hydropathy profile characteristic of receptors with seven putative transmembrane regions. This and the comparison with other cloned receptors indicates that the ADH receptor is a member of the superfamily of G-protein-coupled receptors.

The gene for the peripheral myelin protein PMP-22 is a candidate for Charcot-Marie-Tooth disease type 1A.

Abstract: Charcot-Marie-Tooth disease type 1A (CMT1A) is an autosomal dominant peripheral neuropathy associated with a large DNA duplication on the short arm of human chromosome 17. The trembler (Tr) mouse serves as a model for CMT1A because of phenotypic similarities and because the Tr locus maps to mouse chromosome 11 in a region of conserved synteny with human chromosome 17. Recently, the peripheral myelin gene Pmp-22 was found to carry a point mutation in Tr mice. We have isolated cDNA and genomic clones for human PMP-22. The gene maps to human chromosome 17p11.2-17p12, is expressed at high levels in peripheral nervous tissue and is duplicated, but not disrupted, in CMT1A patients. Thus, we suggest that a gene dosage effect involving PMP-22 is at least partially responsible for the demyelinating neuropathy seen in CMT1A.

Reactive oxygen in skeletal muscle. I. Intracellular oxidant kinetics and fatigue in vitro.

Abstract: We hypothesized that muscle fiber bundles produce reactive oxygen intermediates and that reactive oxidant species contribute to muscular fatigue in vitro. Fiber bundles from rat diaphragm were moun...

Crystallographic evidence of a large ligand-induced hinge-twist motion between the two domains of the maltodextrin binding protein involved in active transport and chemotaxis.

Abstract: The periplasmic maltodextrin binding protein of Escherichia coli serves as an initial receptor for the active transport of and chemotaxis toward maltooligosaccharides. The three-dimensional structure of the binding protein complexed with maltose has been previously reported [Spurlino, J. C., Lu, G.-Y., & Quiocho, F. A. (1991) J. Biol. Chem. 266, 5202-5219]. Here we report the structure of the unliganded form of the binding protein refined to 1.8-A resolution. This structure, combined with that for the liganded form, provides the first crystallographic evidence that a major ligand-induced conformational change occurs in a periplasmic binding protein. The unliganded structure shows a rigid-body "hinge-bending" between the two globular domains by approximately 35 degrees, relative to the maltose-bound structure, opening the sugar binding site groove located between the two domains. In addition, there is an 8 degrees twist of one domain relative to the other domain. The conformational changes observed between this structure and the maltose-bound structure are consistent with current models of maltose/maltodextrin transport and maltose chemotaxis and solidify a mechanism for receptor differentiation between the ligand-free and ligand-bound forms in signal transduction.

Two independent mutational events in the loss of urate oxidase during hominoid evolution.

Abstract: Urate oxidase was lost in hominoids during primate evolution. The mechanism and biological reason for this loss remain unknown. In an attempt to address these questions, we analyzed the sequence of urate oxidase genes from four species of hominoids: human (Homo sapiens), chimpanzee (Pan troglodytes), orangutan (Pongo pygmaeus), and gibbon (Hylobates). Two nonsense mutations at codon positions 33 and 187 and an aberrant splice site were found in the human gene. These three deleterious mutations were also identified in the chimpanzee. The nonsense mutation at codon 33 was observed in the orangutan urate oxidase gene. None of the three mutations was present in the gibbon; in contrast, a 13-bp deletion was identified that disrupted the gibbon urate oxidase reading frame. These results suggest that the loss of urate oxidase during the evolution of hominoids could be caused by two independent events after the divergence of the gibbon lineage; the nonsense mutation at codon position 33 resulted in the loss of urate oxidase activity in the human, chimpanzee, and orangutan, whereas the 13-bp deletion was responsible for the urate oxidase deficiency in the gibbon. Because the disruption of a functional gene by independent events in two different evolutionary lineages is unlikely to occur on a chance basis, our data favor the hypothesis that the loss of urate oxidase may have evolutionary advantages.